1. Field of the Invention
This invention relates to an apparatus for extracting magnetic particles suspended in a fluid, and particularly for sedimenting and concentrating immunomagnetic particles for analysis.
2. Description of Prior Art
The general technology of using antibody-coated magnetic beads or other magnetic particles, hereinafter referred to as immunomagnetic particles or IMPs, to selectively separate and capture analytes from foods or other samples is known as immunomagnetic separation (IMS) and is widely used. In a typical IMS procedure IMPs are suspended in a suspension of the test sample for a time sufficient for them to selectively bind the target analyte and are then pulled out of the suspension as a small pellet-like sediment by means of a strong magnet. After pouring or pipetting away the supernatant suspension the IMPs can be rinsed by resuspending the pellet in clean diluent and resedimenting it with the magnet, after which the target analyte-bearing IMPs can be introduced into whatever final procedure has been chosen to detect or quantify the target analyte. As each sedimentation usually requires only seconds, capture by IMPs is a convenient and rapid first step in many analyses.
Many different systems and individual pieces of apparatus have been developed to assist the use of IMS. A strong magnet is required in order to maximise the speed with which IMPs can be drawn down and generally the magnets used in these systems are of the Neodymium-Iron-Boron alloy type, commonly referred to as “rare earth magnets”. The sedimenting force acting on an IMP at any point in the suspension depends on the magnetic flux density at that point, and because this usually decreases very rapidly with increasing distance from the surface of a magnet, IMP systems are generally designed to handle small volumes, for example 1-10 mL, in small tubes such as the Eppendorf or similar-sized centrifuge tubes commonly found in analytical laboratories.
This small volume results in a limit on the detectable quantity or concentration of target analyte that is often too high for the requirements. For example, if a regulatory standard demands that Listeria monocytogenes must not be detectable in a 25 g sample of food then an acceptable Listeria detection procedure must be capable of detecting the presence of even a single cell of the bacterium in a sample. However as no known technique can detect such a target analyte until it has been removed from the test sample into liquid suspension the first step in any analysis would be to shake or blend the 25 g sample with 225 mL of sterile diluent. The single cell the analysis must detect may now be anywhere in the 250 mL volume of sample-plus-diluent and the probability of capturing it in even a 10 mL aliquot by means of IMPs will be unacceptably low. Without means to treat the whole 250 mL suspension by IMS the only recourse is to incubate the suspension for hours or days to allow the target cells to multiply to a high enough concentration that the aliquot has a reasonable probability of containing target cells. This time delay is a serious impediment to rapid analysis.
A commercially available system for capturing microorganisms from 250 mL volumes using IMPs (Pathatrix™, Matrix MicroScience Limited, Lynxx Business Park, Fordham Rd, Newmarket, UK), comprises a set of peristaltic pumps, vessels, tubes and in-line filters. The magnet and IMP-capturing dimensions of this system are essentially similar to those used in the small-tube apparatus described above and the system is able to handle the larger volume by pumping the suspension slowly and continually past the magnet. This requires time, although protocols for using the Pathatrix™ system may include time for multiplication of a target microorganism. Assembling its tubes and vessels and removing the captured IMP pellet for introduction to the detection step of an analysis is inconvenient and time consuming. Therefore, it would be desirable to have a simpler and faster means to rapidly capture IMPs from large volumes without need for tubes and pumps.
In my previous Canadian patent application, CA 2685229 to C. I. Bin Kingombe and A. N. Sharpe, it is disclosed that it is possible first to sediment IMPs from 250 mL of suspension to the base of a 500 mL glass Erlenmeyer (conical) flask by standing the flask over a powerful magnet and then to induce them to concentrate to a “pellet” at the centre of the base by intensely vibrating the flask axially at high frequency over a second magnet assembly arranged to produce a magnetic field radiating horizontally from the centre of the base. This vigorous vibration is required to overcome stiction of IMPs against the glass. While this device may be useful in a research laboratory, it has numerous shortcomings that make it quite unsuited for routine use in analytical laboratories. For example the concave conical flask base makes it difficult for a motivating magnetic field situated beneath the flask to persuade IMPs to move “uphill” so that it is necessary for the apparatus to vibrate noisily for periods of up to ten minutes whilst sedimented IMPs coalesce at the centre of the base. Moreover once a pellet of IMPs has formed it is not easily removed for analysis owing to the height of a conical flask and it is necessary to modify for example a “magnetic pipet” such as the commercially available PickPen™ product (Bio-Nobile Oy, Tykistokatu 4B, Turku 20521, Finland) in order to make it long enough to reach the bottom of the flask. Furthermore, as it simply rests on the summit of the curved base of the flask without any form of physical restraint the pellet is easily disturbed. Additionally it is not possible to see the pellet if the suspension is cloudy and as the inner flask base slopes away from the centre and glass is relatively slippery it is entirely unable to help the user by passively guiding the point of the pipet into the pellet and it was necessary to include a system whereby the pelletising magnet swings away to reveal a mirror by which the user can see both the pellet in the centre of the base and the tip of the pipet without bending over to peer upwards from beneath.